1. Field of the Invention
The present invention relates to a secretory expression vector capable of extracellularly secreting heterologous polypeptides in mycobacteria.
2. Discussion of the Background
BCG, which is one of the mycobacteria, is an attenuated strain obtained by subculturing bovine tubercle bacillus selected among tubercle bacilli capable of proliferating even in macrophage in an artificial medium over 20 years. BCG is characterized in that it is the only live vaccine which is currently being used as a bacterial vaccine in human. It has extremely weak toxicity, and it has a strong non-specific cellular immunity potentiating effect (adjuvant activity) not only in tuberculous infection but also in various other infections. It is also relatively inexpensive. Accordingly, if BCG can be made to secrete antigenic polypeptides of various pathogenic bacteria or viruses by using genetic engineering techniques, it would be possible to develop a live vaccine of recombinant BCG having the following advantages:
(1) high safety; PA1 (2) long-lasting activity; PA1 (3) vaccine activity induced by the strong adjuvant activity of BCG per se; PA1 (4) low costs;
At present, the vaccinia virus has been widely used as a vector in the study of recombinant live vaccines. However, the vaccinia virus has drawbacks because there is the danger that a virulent revertant might appear. For example, encephalitis might be caused when inoculated in humans. Thus, the vaccinia virus is far from practical use and attention has now been directed to BCG, which is safe and can provide a strong adjuvant effect.
On the other hand, no report of the expression of exogenous genes in mycobacteria by genetic engineering has been made since the transformation system itself has not been established yet. However, this goal has recently become possible because the transformation system by electroporation has been established. (Proc. Natl. Acad. Sci. U.S.A., 85, 6987 (1988)). In this system, a shuttle vector of Escherichia coli-mycobacteria fused with vector pIJ666 of Escherichia coli and plasmid pAL5000 derived from Mycobacterium fortuitum (FEMS Microbiol. Lett., 30, 221 (1985).) is introduced into mycobacteria by electroporation. Using this system, it was shown that a kanamycin-resistance gene (kanamycin phosphotransferase gene), derived from Escherichia coli, can be expressed in mycobacteria.
However, no systems capable of secreting exogenous polypeptides from mycobacteria such as proteins of the AIDS virus are available in the prior art. Such a system would provide a more effective vaccine.